Mitochondrial proton cycling is responsible for a significant proportion of basal or standard metabolic rate, so further uncoupling of mitochondria may be a good way to increase energy expenditure and represents a good pharmacological target for the treatment of obesity. Uncoupling by 2,4-dinitrophenol has been used in this way in the past with notable success, and some of the effects of thyroid hormone treatment to induce weight loss may also be due to uncoupling. Diet can alter the pattern of phospholipid fatty acyl groups in the mitochondrial membrane, and this may be a route to uncoupling in vivo. Energy expenditure can be increased by stimulating the activity of uncoupling protein 1 (UCP1) in brown adipocytes either directly or through beta 3-adrenoceptor agonists. UCP2 in a number of tissues, UCP3 in skeletal muscle and the adenine nucleotide translocase have also been proposed as possible drug targets. Specific uncoupling of muscle or brown adipocyte mitochondria remains an attractive target for the development of antiobesity drugs.

The interaction of azole antifungal antibiotics with purified Candida albicans cytochrome P-450-dependent 14 alpha-sterol demethylase (P-450DM) was measured spectrophotometrically and by inhibition of enzyme activity. Ketoconazole and ICI 153066 (a triazole derivative) formed low-spin complexes with the ferric cytochrome and induced type II difference spectra. These spectra are indicative of an interaction between the azole moiety and the sixth co-ordination position of P-450DM haem. Both azoles inhibited the binding of CO to the sodium dithionite-reduced ferrous cytochrome, and inhibited reconstituted P-450DM activity by binding to the cytochrome with a one-to-one stoichiometry. Similarly, total inhibition of enzyme activity occurred when equimolar amounts of clotrimazole, miconazole or fluconazole were added to reconstituted P-450DM. These results correlated with the inhibition of P-450DM in broken cell preparations, confirming that all five azoles are potent inhibitors of ergosterol biosynthesis in C. albicans.

Western blots detected uncoupling protein 3 (UCP3) in skeletalmuscle mitochondria from wild-type but not UCP3 knock-out mice. Calibration with purified recombinant UCP3 showed that mouse and rat skeletal muscle contained 0.14 µg of UCP3\mg of mitochondrial protein. This very low UCP3 content is 200-700-fold less than the concentration of UCP1 in brown-adiposetissue mitochondria from warm-adapted hamster (24-84 µg of UCP1\mg of mitochondrial protein). UCP3 was present in brown-adipose-tissue mitochondria from warm-adapted rats but was undetectable in rat heart mitochondria. We expressed human UCP3 in yeast mitochondria at levels similar to, double and 7-fold those found in rodent skeletal-muscle mitochondria. Yeast mitochondria containing UCP3 were more uncoupled than empty-vector controls, particularly at concentrations that were 7-fold physiological. However, uncoupling by UCP3 was not

This study supports our previous work demonstrating weight gain and increased feeding behavior induced by olanzapine and could have important implications for enhancing our understanding of the mechanisms by which olanzapine and other atypical antipsychotics induce weight gain in the clinic.

Candida albicans chitinase isolated using the Dyno-Mill disruption technique was characterized using an improved radiometric assay procedure. The enzyme had apparent temperature and pH optima of 45 "C and 6.5, respectively. The preparation yielded an apparent K , of 3.9 mg chitin ml-1 [ 17.6 mM-N-acetylglucosamine (GlcNAc) equivalents] and V of 2.3 nmol GlcNAc formed min-l (mg protein)-'. The potential of the streptomycete antibiotic allosamidin as an antifungal agent is discussed in view of its dose-dependent inhibition of C. albicans chitinase activity (ICs0 = 0.3 VM). Allosamidin was a potent competitive inhibitor of enzyme activity (Ki = 0-23 VM).